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Role of NADPH-insensitive nitroreductase gene to metronidazole resistance of Helicobacter pylori strains
M Kargar,M Baghernejad,A Doosti
DARU : Journal of Pharmaceutical Sciences , 2010,
Abstract: Background and the purpose of the study: Current anti-H. pylori therapies are based on the use of two antibiotics with a proton pump inhibitor and/or a bismuth component. Metronidazole is a key component of such combination therapies in Iran. The aim of this study was to determine the role of rdxA gene in resistant strains of H. pylori isolated from Shahrekord Hajar hospital to metronidazole. Methods: This study was a cross-sectional method, which was carried out on 263 patients who referred to endoscopy department of Hajar hospital, in 2007. Biopsy samples were cultured on selective Brucella agar containing 10% blood and incubated under microerophilic condition at 370C for 3 - 7 days. Suspected colonies were tested by Gram staining, urease, oxidase and catalase activities. Organisms were confirmed to be H. pylori on the basis of the presence of ureC(glmM) gene by PCR .Specific primers were used for detection of rdxA gene mutation . Results: Eighty and four strains of H. pylori determined by PCR method. Of the isolated strains, 49 (58.33%) were resistant, 7 (8.33%) were semi-sensitive to metronidazole and 200bp deletion in rdxA gene was observed in 2 strains. Conclusion: Because of the high metronidazole resistance in patients under study it was necessary to replace it by other antibiotics in therapeutic regimens. On the basis of low frequency of resistance mutation in rdxA gene, sequence analysis for identification of other mechanisms is suggested.
Preparative SDS-PAGE Electroelution for Rapid Purification of Alkyl Hydroperoxide Reductase from Helicobacter pylori
T Mohammadian,M Doosti,M Paknejad,F Siavoshi
Iranian Journal of Public Health , 2010,
Abstract: "nBackground: Alkyl hydroperoxide reductase (AhpC) of Helicobacter pylori is considered as a diagnostic antigen. There-fore, this antigen can be used to detect H. pylori infection by stool immunoassays such as ELISA. The aim of this study was to simplify the AhpC protein purification procedures."nMethods: For whole cell protein extraction, the bacterial cells were ruptured by octly-β-D glucopyranoside. The isolation and purification of AhpC protein were attempted by various techniques including ammonium sulfate precipitation, dialysis, preparative sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and electroelution."nResults:A simple method was used for protein purification AhpC protein. One-dimensional preparative gel electrophoresis allows a single and short purification step; the high resolution capacity of this technique leads to a high level of purity of the protein. Moreover, it avoids contamination by other non-specific proteins which often appear during protein purification by column chromatography."nConclusion: The present method is simple, rapid and makes it possible to preparate AhpC from H. pylori
The effect of Alcoholic garlic (Allium sativum) extract on ABCA1 expression in human THP-1 macrophages
Malekpour-Dehkordi Z,Javadi E,Doosti M,Paknejad M
Tehran University Medical Journal , 2011,
Abstract: "n Normal 0 false false false EN-US X-NONE AR-SA MicrosoftInternetExplorer4 /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Table Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-qformat:yes; mso-style-parent:""; mso-padding-alt:0in 5.4pt 0in 5.4pt; mso-para-margin:0in; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:11.0pt; font-family:"Calibri","sans-serif"; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-fareast-font-family:"Times New Roman"; mso-fareast-theme-font:minor-fareast; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin; mso-bidi-font-family:Arial; mso-bidi-theme-font:minor-bidi;} Background: ATP-binding cassette transporter A1 (ABCA1) is a key mediator of cholesterol efflux to apoA-I in lipid-laden macrophages, the first step of reverse cholesterol transport (RCT) in vivo and a critical step in preventing atherosclerosis. Enhanced ABCA1 expression may inhibit foam cell formation and consequently reduce atherogenic risk. On the other hand, garlic, Allium sativum, and garlic extracts have been demonstrated to have potential cardiovascular benefits. Moreover, garlic has direct antiatherogenic and antiathersclerotic effects on artery walls. The aim of this study was to evaluate the effects of alcoholic garlic extract on the expression of ABCA1 in macrophages."n"nMethods: Cell viability assay was used in order to detect the cytotoxic dose of alcoholic garlic extract on macrophages. Real-time PCR and Western blotting were performed to study the effects of alcoholic garlic extract on the expression of ABCA1. Macrophage cells were treated by different concentrations of alcoholic garlic extract for 48 h. The total RNA of the treated macrophages were extracted and analyzed by real-time PCR. ABCA1 protein expression was also analyzed using the Western blotting technique."n"nResults: Alcoholic garlic extract increased the ABCA1 mRNA (20-23%) and protein expression (18-37%) in THP-1 macrophage cells compared with the controls (untreated cells)."n"nConclusion: The results of this study are suggestive of the potential effects of alcoholic garlic extract in increasing ABCA1 expression in macrophages, the possibility of promoting reverse cholesterol efflux in macrophages and preventing atherosclerosis.
Determining Effects of Elaidic Acid on PPAR- Gamma Expression in RAW 264.7 Macrophage Cell Line
H Montakhab Yegane,H Babaahmadi Rezaiy,M Doosti
Tehran University Medical Journal , 2012,
Abstract: Background: Several dietary factors are involved in cardiovascular coronary heart diseases, including trans fatty acids, which are generally formed during hydrogenation of vegetable oils, a process that causes conversion of liquid oils into semisolid fats. Nowadays, it is well-known that trans fatty acids form a major risk factor in the occurrence and progression of atherosclerosis. On the other hand, it has been identified that some nuclear receptors, such as PPARs, are involved and play important roles in lipid homeostasis and pathogenesis of cardiovascular diseases. Therefore, we studied the effect of elaidic acid on gene expression of peroxisome proliferator activated receptor gamma (PPARγ).Methods: Murine macrophage RAW264.7 cells were treated by 0.5, 1, and 2 mM concentrations of elaidic acid for 6 h. The control group was treated by 50% ethanol (as solvent), equivalent to the amount of ethanol used in 2 mM concentration of elaidic acid. Later, the total RNA was extracted and its cDNA was synthesized. Finally, the quantity of PPARγ gene expression was measured by real-time PCR.Results: Overall, 0.5, 1, and 2 mM concentrations of elaidic acid decreased PPARγ gene expression in RAW264.7 macrophage cell line by -1.36, -1.68, and -3.24 folds compared with the control group, respectively.Conclusion: By decreasing the expression of nuclear receptor PPARγ, elaidic acid causes, intensifies or accelerates the occurrence of cardiovascular diseases, especially atherosclerosis. This finding shows the importance of reducing the consumption of elaidic acid containing foods.
Mitochondrial DNA (mtDNA) structure of Anopheles superpictus populations in IRAN
Zavarreza J,Doosti M,Ariabarzin Sh,Soleymani S
Tehran University Medical Journal , 2007,
Abstract: Background: Malaria is still one of the main health problems in south and southeast provinces of Iran and recently on average 10,000-30,000 malaria cases were reported annually. Mosquitoes of Anopheles superpictus are one of the main malaria vectors in Iran and have been reported from all areas of the country including central plateau and plains of Alborz and Zagrous Mountains chains, and with low numbers in shore plains of the Persian Gulf and Caspian Sea. There are variations in larval and adult morphological characters and also in vectorial capacity of this species in different areas of Iran. Methods: This study has been conducted to investigate rate of mtDNA variation among various populations of this species in Iran. The sequence variation of an 1512 bp length of mitochondrial DNA (mtDNA) cytochrome oxidase subunits 1 and 2 (COI-COII) and an 708 bp sequences of COI gene were analyzed by PCR-RFLP and PCR-direct sequencing respectively. Results: This study showed that there are considerable variations between and within populations. Rate of variation was 12.3 % between populations and this was 2-5% for within Baluchistan population. Totally 4 haplotypes were observed between populations where 3 occur in Baluchistan and one in other places. Conclusion: This is the first report on existence of various haplotypes in An. superpictus in science, and presumably this species comprising siblings and is a species complex. Further studies need to confirm this result and to determine the relationship between mtDNA haplotypes and their role in malaria transmission in each locality.
Extraction and separation of lipopolysaccharide from outer membrane of Helicobacter pylori
Zavarreza J,Doosti M,Ariabarzin Sh,Soleymani S
Tehran University Medical Journal , 2007,
Abstract: Background: Helicobacter pylori (H. pylori) is one of the major causes of peptic ulcer, gastritis and gastric cancer. This bacterium has a special lipopolysaccharide (LPS), which is responsible for its pathogenesis and its high resistance against gastric acid and escape from the human immune system. This property makes it a target for further research and diagnostic goals. In this study, the extraction of the LPS and separation from the outer membrane is required. Methods: The LPS was extracted from the outer membrane, or envelope, of H. pylori obtained from patients suffering from gastritis, gastric ulcer and gastric cancer. LPS extraction was performed using the proteinase K method. SDS-PAGE and silver staining were applied to investigate the electrophoretic pattern of the LPS. This pattern was compared with that of E. coli serotype O111:B4 and Salmonella serotype ATCC 14028. Results: The extracted LPS has a ladder-shaped electrophoretic pattern and the bands are located in three groups: high, medium and low molecular weights. Conclusion: The distribution of the bands of the ladder-shaped electrophoretic pattern is caused by the different number of oligosaccharide chains associated with the LPS. The high molecular weight bands represent S-LPS and the low molecular weight bands represent the R-LPS, which lacks the O-chain.
Evaluation of antioxidant status in Streptozotocin-induced diabetic rats by Ferric reducing ability of plasma assay
Zarban A,Doosti M,Esteghamati A,Allameh A
Acta Medica Iranica , 2000,
Abstract: Enhanced oxidative stress in diabetes mellitus may contribute to the pathogenesis of diaetic complications. In this article, the stress oxidative generation has been studied in experimental diabetes by ferric reducing ability of plasma assay, a sensitive and simple method, and by other oxidative damage markers. The ferric reducing ability of plasma values as a total antioxidant capacity were significantly decreased at the 3th 4th weeks of stuy ( P<0.02), while the thiobarbituric acid reacting substances (TBARS) levels in plasma wre increased at the 3rd and the 4 th weeks ( (P<0.05). the plasma carbonyl groups (PCG) were not affected, and total thiol groups (TG) were significantly decreased at the 4th week (P<0.02).In conclusion the present study suggests that hyperglycemia in diabetes leads to oxidative stress, as shown by ferric reducting ability of plasma assay. This method is rapid, simple and economic
ASSOCIATION BETWEEN THE SEVERITY OF ANGIOGRAPHIC CORONARY ARTERY DISEASE AND PARAOXONASE-1 PROMOTER GENE POLYMORPHISM T(-107)C IN IRANIAN POPULATION
A. Jalilian,E. Javadi,M. Doosti,P. Amiri
Acta Medica Iranica , 2008,
Abstract: The oxidation of low-density lipoproteins and cell membrane lipids is believed to play an integral role in the development of fatty streak lesions, an initial step in coronary artery disease (CAD). Paraoxonase-1 (PON1) is an enzyme associated with the high-density lipoprotein (HDL) particle. PON1 protects LDL from oxidative modification by hydrolyzing lipid peroxides, suggestive of a role for PON1 in the development of CAD. The present study tested the hypothesis that Paraoxonase-1 promoter polymorphism T(-107)C could be a risk factor for severity of CAD in Iranian population. Paraoxonase-1 promoter genotypes were determined in 300 consecutive subjects (> 40 years old) who underwent coronary angiography (150 subjects with >50% stenosis served as cases [CAD+] and 150 subjects with < 20% stenosis served as controls [CAD-]). PON1 promoter genotypes were determined by PCR and BSTU1 restriction enzyme digestion. CAD+ Subjects did not show any significant differences in the distribution of PON1 promoter genotypes as compared to CAD- Subjects (P = 0.075). However the analysis of PON1 promoter genotypes distribution showed a higher percentage of (-107) TT among CAD+ compared with CAD- (P = 0.027). After controlling for other risk factors, the T(-107)C polymorphism had interaction with age (P = 0.012), but did not show any interaction with other risk factors such as BMI ,gender, smoking, diabetes, level of HDL-C, LDL-C, triglyceride and Total cholesterol. These data suggest that the TT genotype may represent a genetic risk factor for Coronary artery disease in Iranian population.
"APO(a) isoforms and LP(a) concentration in predicting risk for coronary artery disease: A study in men <55 years of age "
"Rashtchizadeh N,Javadi E,Doosti M,Mohagheghi A
Acta Medica Iranica , 2000,
Abstract: Lipoprotein (a) [Lp (a) is formed by assembly of LDL-particles and the carbohydrate rich protein, apolipoprotein (a) [apo (a)]. Elevated plasma Lp (a) levels are an independent predictor of the development of premature coronary artery disease (CAD), but is not clear whether the apo (a) isoform plays on additional and independent role or not. To investigate the possible effect of apo (a) isoform on premature CAD (in patients < 55 years of age), we have analyzed apo (a) isoforms, Lp (a) level and their relation with many recognized CAD risk factors, in 60 male patients with angiographically defined CAD and in 60 male control with no angiographic evidence of CAD. The results show elevated Lp (a) concentration (29.4±16.1, vs. 16.5±9.9 P<0.01) and frequency of S2 isoform (31.7%, vs. 6.7% P<0.01) and B isoform (10% vs. 1.7% P<0.01) in patients with premature CAD. Patients with S2 isoform exhibited significantly higher plasma Lp(a) concentration than control subject with the same isoform (39.8±15.9 vs. 20.5±6.9, P<0.05), but patient with B isoform exhibited no significant Lp(a) concentration as compared to the controls (49.5±9.46 vs. 45). In addition, all patients had a low frequency of S4 and null isoforms. The distribution of apo (a) lsoforms was significantly shifted towards small isoform size (band S2) in the CAD as compared to the controls. This study provides evidence that CAD patient < 55 years of age have a different pattern of apo (a) isoforms than controls, and therefore apo (a) isoform may play an important role in predicting premature CAD.
Autoantibodies against modified low density Lipoprotein in patients with coronary heart disease and normal individuals
"Mohammadi R,Doosti M,Javadi E,Sarrafnejad A
Acta Medica Iranica , 2001,
Abstract: Different classes of autoantibodies against two antigenic forms of modified low density lipoprotein (mod-LDL) were detected in 140 patients by indirect ELISa method. Investigated autoantibodes included total immunoglobulins (IgT). Immunoglobulin G (IgG), and immunoglobulin M (IgM) against oxidized LDL with copper ions (ox-LDL) and modified LDL with malondialdehyde (mal-LDL) (IgT-O) IgT against mal-LDL (IgT-M), IgG against ox-LDL (IgG-O), IgM against ox-LDL (IgM-o), and IgM against mal-LDL (IgM-M) were significantly (P<0.05) higher in patient with coronary atherosclerosis than normal individuals, whereas titer of IgG against mal-LDl (IgG-M) didn’t show any significan difference between these groups. In this study, no correlation was found between autoantibody titers and severity of coronary artery stenosis. The results indicate that titers of these autoantibodies are dependent on an active atherogenic process.
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